FBXO10 deficiency and BTK activation upregulate BCL2 expression in mantle cell lymphoma

Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to identify BTK targets by RNA-seq and high-throughput data analysis and verify these genes by quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods Methods: mino/z138 sc4/shbtk stable cell lines were generated with tet-on system vector, small hairpins were induced for 48 hours after doxycycline addition, mRNA was exacted and used for RNA sequencing. After TopHat analysis followed by Cufflinks, down/up regulated genes list was generated. qRT–PCR validation was then performed using SYBR Green assays Results: Using an optimized data analysis workflow and with a fold change =1.3 and p value <0.05, thousands of genes were changed. Altered expression of 6 genes was then confirmed with qRT–PCR, demonstrating the high qulity of the RNA-seq method. Overall design: mino/z138 sc4/shbtk stable cell lines were generated with tet-on system vector, small hairpins were induced for 48 hours after doxycycline addition, total RNA from these cells was then used for RNA-seq analysis.